Method for Hemostasis Testing

ABSTRACT

A sample testing cartridge is usable to perform a variety of tests on a viscoelastic sample, such hemostasis testing on a whole blood or blood component sample. The cartridge includes a sample processing portion that is in fluid communication with a sample retention structure. A suspension, such as a beam, arm, cantilever or similar structure supports or suspends the sample retention portion relative to the sample processing portion in a unitary structure. In this manner, the sample retention portion may be placed into dynamic excitation responsive to excitation of the cartridge and correspondingly dynamic, resonant excitation of the sample contained within the sample retention portion, while the sample processing portion remains fixed. Observation of the excited sample yields data indicative of hemostasis. The data may correspond to hemostasis parameters such as time to initial clot formation, rate of clot formation, maximum clot strength and degree of clot lysis.

CROSS-REFERENCE TO RELATED APPLICATION

This patent claims priority to U.S. Provisional Patent Application Ser.No. 61/729,349, filed Mar. 15, 2014 entitled Apparatus, Cartridge andMethod for Hemostasis Testing the disclosure of which is herebyexpressly incorporated herein by reference.

STATEMENT AS TO RIGHTS TO INVENTIONS MADE UNDER FEDERALLY SPONSOREDRESEARCH OR DEVELOPMENT

This invention was made with government support under grants R43HL088850and R44HL088850, awarded by the National Heart, Lung, and BloodInstitute of the National Institutes of Health. The U.S. Government hascertain rights to this invention.

TECHNICAL FIELD

This patent relates to blood hemostasis testing, and in particular, thispatent relates to sample testing cartridges for preparing and testingblood samples.

BACKGROUND

Blood is in liquid form when traveling undisturbed in bodilypassageways. However, an injury may cause rapid clotting of the blood atthe site of the injury to initially stop the bleeding, and thereafter,to help in the healing process. An accurate measurement of the abilityof a patient's blood to coagulate in a timely and effective fashion andsubsequently to lyse is crucial to certain surgical and medicalprocedures. Also, accurate detection of abnormal hemostasis is ofparticular importance with respect to appropriate treatment to be givento patients suffering from clotting disorders.

Blood hemostasis is a result of highly complex biochemical processesthat transform the blood from a liquid state to a gel state.Characteristics of blood, such as strength of the clot and othermechanical properties of the blood are useful in determining itshemostasis characteristics. For example, if the strength of the clot canresist the shear forces of the circulating blood, that clot can adhereto a damaged vascular site (e.g. open vascular system following surgery)and stop bleeding. That same formed clot in an undamaged (i.e. closed)vascular system will impede the flow of blood and, depending on itslocation, can cause heart attack, ischemic stroke, pulmonary embolism(PE), or deep vein thrombosis (DVT).

In accordance with commonly owned U.S. Pat. No. 8,236,568 entitledMethod of Analyzing Hemostasis; U.S. Pat. No. 7,879,615 entitledHemostasis Analyzer and Method and U.S. Pat. No. 7,261,861 entitledHemostasis Analyzer and Method, the disclosures of which are herebyexpressly incorporated by reference, a description is provided ofapparatus and methods for hemostasis analysis by observation of sampleresonant response to dynamic excitation. A blood hemostasis analyzer inaccordance with the teachings of these patents operates under theprinciple that because hemostasis of a blood sample changes the bloodsample from a liquid state to a gel state, and the viscoelasticproperties of the clot formed by coagulation controls the naturalfrequency of the sample, measuring changes in the natural frequency ofthe blood sample during coagulation provides the hemostasischaracteristics of the blood sample. In keeping with this principle, theblood hemostasis analyzer measures the changes in the natural frequencyof a blood sample during coagulation and lysis to provide hemostasischaracteristics of the blood sample. To measure hemostasis in thismanner, the analyzer generally includes a container for holding theblood sample, a shaker or exciter for displacing the container to excitethe blood sample to resonant vibration, and a sensor for measuring theresulting amplitude of motion of the blood sample.

The above-patented method of hemostasis analysis provides for vibrationof a sample to resonance. As blood transitions from a liquid state to agel state, such as a substantially dilute cross-linked system,exhibiting no flow at steady-state, the natural frequency of the bloodsample increases. Hence, measuring the changes in the natural frequencyof the sample under excitation and during clotting and lysis provides ahemostasis indication.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 graphically depicts a cartridge for sample testing in accordancewith a herein described embodiment of the invention.

FIG. 2 is a schematic illustration of a hemostasis analyzer within whichthe cartridge of FIG. 1 may be used.

FIG. 3 graphically depicts a cartridge for sample testing in accordancewith a herein described alternate embodiment of the invention.

FIG. 4 is a front view of a sample containment structure according to aherein described embodiment.

FIG. 5 is an end view of the sample containment structure shown in FIG.4.

FIG. 6 is a front view of a sample containment structure according to aherein described embodiment.

FIG. 7 graphically depicts a cartridge for sample testing in accordancewith a herein described embodiment of the invention.

SUMMARY

A cartridge for sample testing may include a sample preparation portionincluding a fluid processing structure and a sample testing portion thatincludes a sample retention structure. The sample testing portion may becoupled to the sample preparation portion by a fluidic passage providingfluid communication between the fluid processing portion and the sampleretention structure. The sample retention structure supports a sample tobe tested such that the sample may be excited to resonant vibrationresponsive to an excitation applied to the cartridge. The sampleretention structure also permits observation of the sample excited toresonant vibration.

The cartridge may be coupled to the sample preparation portion such thatthe sample retention structure may be excited separate and apart fromthe sample preparation portion. For example, the sample retentionstructure may be coupled via a suspension to the sample preparationportion.

The cartridge may be an assembly of components. For example, thecartridge may be a laminate structure comprising a number of separatelayers joined together.

The fluid processing structure of the cartridge may include operativelyarranged a plurality of fluid passages, at least one valve, a bellowsand a reservoir. A reagent may be disposed in any one or more of thefluid passage, valve, bellows, reservoir or other portions of the fluidprocessing structure and multiple combinations thereof.

A valve structure of the cartridge may be a hydrophobic vent surfacedisposed within a passage or other fluidic structure. Another valvestructure may be a flexible membrane disposed within a passage.

The sample retention structure may be an annulus.

A method of testing blood hemostasis may include introducing a bloodsample into a fluid processing structure, where the fluid processingstructure contains a reagent, for example in a passage. The fluid samplemay be processed by passing a portion through the fluid processingstructure so as to contact the reagent. The processed portion is thencommunicated to a sample retention structure of the fluid processingstructure. Testing is accomplished by vibratory excitation of theprocessed portion to obtain data indicative of at least one hemostasisparameter.

The reagent may be disposed within a passage of the fluid processingstructure, and processing the sample may be passing the portion throughthe passage.

The fluid processing structure may include a pump for pumping theportion within the fluid processing structure. The pump may also be usedto pump the sample to the sample retention apparatus.

The fluid processing structure may include first and second passagescorresponding to first and second tests or assays. In such anarrangement, processing the sample may include communicating a firstportion of the blood sample through the first passage and communicatinga second portion of the blood sample through the second passage.Likewise, the first and second portions may be communicated to first andsecond sample retention structures. Simultaneous testing of multipleprocessed sample portions may be accomplished, for example by excitingthe processed portion or portions to resonant vibration. Sampleexcitation may be accomplished by applying an excitation signal to thecartridge containing the fluid processing structure and the sampleretention structure, or by applying an excitation signal only to aportion of the cartridge containing the sample retention structure.

Generated hemostasis data may include parameters at least indicative ofone of: time to initial clot formation, rate of clot formation, maximumclot strength and time to clot lysis and degree of clot lysis at apredetermined time after maximum clot strength, e.g., 30 minutes. Thedata may be communicated to other locations or processors over anetwork. The data may also or alternatively be depicted graphically.

DETAILED DESCRIPTION

In accordance with the herein described embodiments, a sample-testingcartridge is usable to perform a variety of tests on a viscoelasticsample, such as a whole blood or blood component sample. The cartridgeincludes a sample-processing portion that is in fluid communication witha sample retention structure. In one embodiment, a suspension, such as abeam, arm, cantilever or similar structure supports or suspends thesample retention portion relative to the sample-processing portion in aunitary structure. The sample-processing portion may be held rigidly soas to permit communication with a source of pneumatic drive as well asuser interaction, while the sample retention portion may be placed intodynamic excitation responsive to excitation of the sample retentionportion of the cartridge, another portion of the cartridge or thecartridge as a whole. Correspondingly, dynamic, resonant excitation ofthe sample is achieved within the sample retention portion as a resultof such excitation. Observation of the excited sample yields dataindicative of the changing elastic properties of the sample. This datamay correspond to hemostasis parameters such as time to initial clotformation, rate of clot formation, maximum clot strength, time to clotlysis and degree of clot lysis.

FIG. 1 graphically depicts a sample testing cartridge 10 that includes asample processing portion 12, a sample retention portion 14 and asuspension, e.g., beam 16 structurally, mechanically joining the sampleretention portion 14 to the processing portion 12. The beam 16 shown ina cantilever configuration allows the sample retention portion 14 to actas a sprung mass relative to the sample processing portion 12 and tovibrate in response to a stimulus applied to the cartridge 10. Otherstructures, such as spring, multi-link suspensions, a rigid orsemi-rigid member or members and the like capable of mechanicallyjoining while allowing relative, dynamic movement of the sampleretention portion to the sample processing portion may be used. It willbe appreciated relatively small displacement, i.e., vibration of thesample is required. In certain embodiments, it may be possible todirectly join the processing portion 12 and the retention structure 14even forming them as an integral member.

The sample processing portion 12 includes a port 18 through which aliquid sample 100 may be introduced into the sample processing portion12. The port 18 may be self-sealing (as in a septum or other automaticsealing mechanism) such that the sample once introduced into thecartridge 10 does not flow, leak, seep, etc. from the cartridge. Theport 18 communicates with a reservoir 20 into which the sample isinitially received. The sample processing portion 12 additionallyincludes channels, via, waste chambers, passages and similar structures22; a bellows or pump 24 and valves 26 to control movement of the sample100 or a portion thereof through the sample processing portion 12responsive to actuation of the bellows 24 to prepare the sample 100 fortesting.

Pneumatic force, which can be applied pressure, drawn vacuum orcombinations thereof, and in a preferred implementation is vacuum, maybe used directly on the sample 100 to move it into the cartridge 10 andto manipulate the various elements of the cartridge 10. In theillustrated implementation, vacuum applied at a central port 19 causesthe sample 100 to load into a staging area 20 and further draws thesample 100 into the bellows 24. The sample 100 is drawn up to thehydrophobic vents 28, allowing careful control of the sample fluidvolumes solely with the card geometry. As such, it is unnecessary tomonitor loading time or otherwise actively sense of the volume of thesample 100, simplifying the structure and operation of the cartridge 10.

Application of vacuum to the bellows 24 and operation of selected valves26 causes the sample portion 100 to be drawn from the staging area 20and through and into a first passage 22′. The first passage 22′ mayinclude a testing reagent, in liquid, gel, lyophilized, dried or othersuitable form that is reconstituted by, and then mixed with the sampleportion 100 as it is drawn into and through the passage 22′. Cycling ofthe bellows 24 provides mixing of the sample and reagent by repeatedcommunication of the sample 100 into and through the first passage 22′.Control of the valve 26 and actuation of the bellows 24 then allowscommunication of the conditioned sample portion 100 through a secondpassage 22″ to the sample retention structure 14.

Bellows 24 operation to communicate the sample portion 100 through thecartridge 10 is not limited to operating the bellows in a binaryfashion. Applying pneumatic pressure and/or vacuum to the bellow 24 viapredetermined profiles, for example ramps, arcs and the like, provides avery controlled approach to the fluid flow profile within the sampleprocessing portion 12 to limit fluid shear in the passages 22 which canlead to sample activation and furthermore to avoid bubble formation.Pneumatic inputs to the cartridge 10 and the bellows 24 through a flowrestriction outside the card filters out pulsations caused bypulse-width-modulation (PWM) operation of the solenoid valve controllingthe bellows 24.

Reagent reconstitution and mixing with the sample 100 may beaccomplished by locating the reagent or multiple reagents at variouslocations within the cartridge 10 and exposing the sample portion 100 tothe reagents. Reagents may be positioned at virtually any otherlocation: wells, passages, via, chambers, bellows, and sample retainers,within the cartridge 10 where the reagents will contact the sampleportion 100. Reagents may further be placed in the sample containmentstructure 30. For example, heparinase may be placed in the staging area20 or other sample reservoir area of the cartridge 10. The sampleportion 100 may then be drawn into the staging area 20 and allowed toremain in contact with heparinase for sufficient time to reconstitutethe dried heparinase and counteract sodium heparin in the sample 100.This is prior to the sample 100 being pulled into the bellows andflowing through a reagent well, i.e., passage 22′, where the treatedsample 100 will contact other reagents. Spot reagents may be appliedvirtually anywhere on the cartridge, and additionally, reagent may coatthe sample containing structure 30. Thus it will be appreciated that acartridge 10 in accordance with various embodiments of the invention mayhave numerous different reagents located at numerous different locationsof the cartridge in virtually any set of combinations.

U.S. Pat. No. 6,613,573 entitled Method and Apparatus for MonitoringAnti-Platelet Agents; U.S. Pat. No. 6,787,363 entitled Method andApparatus for Hemostasis and Blood Management; U.S. Pat. No. 6,797,519entitled Method and Apparatus for Diagnosing Hemostasis; U.S. Pat. No.6,890,299 entitled Method and Apparatus for Monitoring Hemostasis inConnection with Artificial Surfaces Devices; U.S. Pat. No. 7,179,652entitled Protocol for Monitoring Platelet Inhibition; U.S. Pat. No.7,524,670 entitled Protocol for Risk Stratification of Ischemic Eventsand Optimized Individualized Treatment; U.S. Pat. No. 7,811,792 Protocoland Apparatus for Determining Heparin-induced Thrombocytopenia; U.S.Pat. No. 7,939,329 entitled Protocol for Risk Stratification of IschemicEvents and Optimized Individualized Treatment; U.S. Pat. No. 8,008,086entitled Protocol for Monitoring Direct Thrombin Inhibition and U.S.Pat. No. 8,076,144 entitled Protocol for Risk Stratification of IschemicEvents and Optimized Individualized Treatment, the disclosures of whichare hereby expressly incorporated herein by reference, teach a number ofpossible reagents and corresponding assays and protocols. The reagentsmay be as described in these patents or other reagents may be used,and/or the card may be configured to carry out other protocols.

The sample retention structure 14 communicates with the second passage22 and includes a containing structure 30 for holding or containing thesample portion 100 during testing of the sample portion 100. Forexample, the sample retention structure 14 may include an annulus,cylinder, cup, or similar containing structure 30 that provides a samplesurface free to be excited to resonant or near-resonant vibration andobserved by a sensing device. One containing structure 30 includes acontaining wall leaving two surfaces of the sample free to be excited toresonant or near-resonant vibration. The sample may be introduced to thecontaining structure 14 via a side port extending through the containingwall. The above-referenced U.S. Pat. Nos. 8,236,568; 7,879,615 and7,261,861 describe several additional possible sample containingstructures 30, all of which are contemplated suitable structures for usein an embodiment of the cartridge 10.

The cartridge 10 charged with a sample 100 is usable in an apparatus formeasuring hemostasis 102. Depicted schematically in FIG. 2, the elementsof the apparatus 102 are an exciter, shaker or similar stimulusgenerator 104, sensor/detector 106, processor 108, user interface 110and communication link 112. A suitable power supply (not depicted) isprovided. The exciter 104 can be a coil, piezoelectric device, motor,acoustic actuator or any suitable device to cause resonant excitation ofthe sample 100 within the sample retention apparatus 14 via directstimulation of the retention apparatus 14 or indirectly via excitationof the cartridge 10 or a portion of the cartridge 10 or via combinationsthereof. The sensor 106 may be an optical/laser device. The userinterface 110 may be hard buttons, touch screen or any suitableinterface to allow the user to select and initiate a testing protocoland to view or to affect recording or transmitting of the results. Theprocessor 108 operably links these functional elements and facilitatescommunication by the communication link 112, which may be a wireless orwired network interface following any suitable protocol. For example,the communication link 112 may be used to communicate results data to aremote processing facility for analysis and diagnostic interpretationand to receive results analysis for display in data and graphic form viathe user interface 110.

The cartridge 10 is placed within the testing apparatus 102. The bloodsample 100, such as fresh whole blood, blood components, and the like isintroduced into a reservoir 18 within the cartridge 10 via the port 20.The apparatus 102 is configured to selectively apply pneumatic signals,such as drawing a vacuum at a selected position of surface 32 of thecartridge 10 or actuating valves within the cartridge 10, in apredetermined testing protocol to condition the sample portion 100 bymixing with reagent and then communicating it to the sample retentionstructure 14.

The cartridge 10 may be built from laser-cut or die-cut layers laminatedtogether to create functional elements: valves 26, bellows 24, channels22 and fluid holding areas/reservoirs 18. It could also be assembledfrom injection-molded or hot-embossed layers that are then laminated,bonded or otherwise assembled together. Individual layers can beconstructed in a number of ways, depending on when they enter the buildsequence for the card.

The materials for each layer are chosen from suitable materials. Forlaser- and die-cut laminates, structural layers may be of a suitableplastic such as polyethylene terephthalate (PET), biaxially-orientedpolypropylene (BOPP), cyclic olefin polymer (COP) or cyclic olefincopolymer (COC). Laminating adhesive may be provided separately or withstructural layers. Flexible membrane layers, such as used to formvalves, bellows and the like may be polyurethane, silicone,polypropylene (PP) or polyethylene (PE). Conventional conversiontechniques may be used to prepare the layers when they are composed ofmore than one material.

Hydrophobic membrane material and sizes, along with layout of thecartridge, may be chosen to reduce the cost of material used per card,and allow for easier automatic placement of the membranes in thefinished cartridge 10. The size of the pores and thickness of thematerial may be used to control the blood flow rate and volume into thecard and into the bellows. Channels leading to the membranes may be lowvolume to reduce sample or reagent loss. In the instant application,this is done to reduce blood sample loss and prevent errors in reagentconcentration. Other channels may be of higher volume to facilitatesample communication within the card, and for example to the sampleretention structure 14.

In one embodiment, the channel size communicating to the sampleretention structure 14 may be made to be 0.017 square millimeters orgreater in cross-section, to facilitate flow of the processed sample tothe sample retention structure without significant shear activation ofplatelets within the sample or unintended activation as a result of bothshear and exposure to reagent. Suitable dimensions of 0.20 squaremillimeters or greater, and for example, 0.30 square millimeters may beused.

FIG. 3 graphically illustrates a cartridge 200 that may be used insample testing, such as in hemostasis sample testing of a whole blood orblood component sample. The cartridge 200 has features similar to thatof cartridge 10, but provides potential for multiple simultaneous tests.That is, each channel on the cartridge may contain different reagentsand hence constitute different tests or may be configured to provideredundant tests. The combined tests may constitute an assay. Thecartridge 200 is configured to perform up to four (4) testssimultaneously. Although, in use any combination from one (1) to four(4) tests may be performed. Cartridge 200 also demonstrates that acartridge may be made with virtually any number of tests, with FIG. 1and FIG. 3 demonstrating at least single test cartridges and four (4)test cartridges, cartridges of 2 or 3 tests may be made as well ascartridge having more than four (4) tests may be made. A channel orchannels of the cartridge may be a test or tests to test a specificcharacteristic of hemostasis that may be used in an assay of suchhemostasis related characteristics, such as platelet activity, ischemicrisk indicators, or the like as set forth in the afore-mentioned USpatents. The cartridges 200 may be configured to provide multiple testsor may be made to provide the same tests with multiple differentsamples, or combinations thereof.

As seen in FIG. 3, the cartridge 200 is formed with four (4) tests A, B,C and D. Each test on the cartridge 200 includes a sample processingportion 212, a sample retention portion 214 and a suspension, e.g., beam216 structurally, mechanically joining the sample retention portion 214to the processing portion 212. The elements of the respective tests areindicated separately by the alpha designation A, B, C or D. Theplurality of beams 216 shown in cantilever configuration allow thesample retention portion 214 to act as a sprung mass relative to thesample processing portion 212 and to vibrate in response to a stimulusapplied to the retention structure 214 and/or the cartridge 200. Otherstructures, such as spring, multi-link suspensions, a semi-rigidmechanical member or members and the like capable of mechanicallyjoining while allowing relative, dynamic movement of the sampleretention portion to the sample processing portion may be used. It willbe appreciated relatively small displacement, i.e., vibration of thesample is required. In certain embodiments, it may be possible todirectly join the processing portion 212 and the retention structure 214even forming them as an integral member.

The sample-processing portion 212 may include a common port feedingthrough a plenum or manifold or individual ports 218 through which aliquid sample 100 may be introduced into the tests of the sampleprocessing portion 212. The ports 218 may be self-sealing such that thesample once introduced into the cartridge 200 does not flow, leak, seep,etc. from the cartridge. The ports 218 communicate with respectivereservoirs or sample holding areas 220 into which the sample isinitially received. The sample-processing portion 12 additionallyincludes, channels, via, passages and similar structures 222; bellows orpumps 224 and valves 226 to control movement of the sample 100 or aportion thereof through the sample processing portion 212 responsive toactuation of the bellows 224 to prepare the sample 100 for testing.Application of external pneumatic pressure to the bellows 224 andoperation of selected valves 226 causes the sample portion 100 to bedrawn from the reservoir 220 and through and into a first passage 222′and the bellows 224. The first passage 222′ may include a testingreagent, in liquid, gel, lyophilized or dried form that is mixed withthe sample as it is drawn into and through the passage 222′. Asdescribed herein, reagents may be located at other locations of thecartridge 200. Cycling of the bellows 224, which may be accomplished asdescribed by above via pulse-width modulation of the pressure and vacuumsignals, allows mixing of the sample and reagent by repeatedcommunication of the sample 100 into and through the first passage 222′.Control of the valve 226 and actuation of the bellows 224 then allowscommunication of the conditioned sample portion 100 through a secondpassage 22″ to the sample retention structure 214 and sample containmentstructure 230. Suitable waste chambers are provided within the cartridgeto ensure containment of the sample.

The cartridge 200 charged with samples 100 is then prepared and ready tobe introduced into a testing apparatus to perform the tests or tests forwhich the cartridge is configured and to report the respective results.

FIGS. 4 and 5 illustrates a sample retention structure 300 that may beused in any of the cartridges 10 or 200 as the respective sampleretention structures 30 and 230. The sample retention structure 300 hasan annulus body 302 with a first end 304 and a second end 306. The firstend 304 had an end surface 308 within which is formed a relief 310 aboutthe entire circumference. A liquid sample, such as whole blood or bloodcomponents is introduced into the sample retention structure 300, forexample through a side port not depicted, and is driven into the body302 by controlled compression or activation of the bellows 224 towardthe first end 304. The annulus 302 is overfilled to ensure all interiorsurfaces are wetted by the sample, and then the sample volume in theannulus is reduced to form a convex surface 312 (shown in phantom)extending from the first end 302. It is this convex surface 312 that isobserved as the sample within the annulus 302 is set into resonantoscillation as a result of stimulation of the cartridge 10/200. Thediameter of the relief 310 also helps to establish a consistent volumeof sample, along with other geometry of the cartridge and control of thefluid processing structures, to be placed into resonant oscillation, andthus complex calibration processes are not required.

While it is possible to observe the entire convex surface 312, observingonly a portion of the free surface may be sufficient to yield a usableresult. For example, a center portion of the convex surface 312 may beobserved.

FIG. 6 illustrates a sample retention structure 400 that may be used inany of the cartridges 10 or 200 as the respective sample retentionstructures 30 and 230. The sample retention structure 400 has an annulusbody 402 with a first end 404 and a second end 406. The first end 404has an end surface 408 formed to an edge structure 410 about the entirecircumference. The edge structure 410 may be substantially planar, thatis, all points about the edge structure 410 being substantially within acommon plane. A liquid sample, such as whole blood or blood components,is introduced at a side port 412 and is driven into the body 402 bycontrolled compression or activation of the bellows 224 toward the firstend 404. The annulus 402 is overfilled to ensure all interior surfacesare wetted by the sample, and then the sample volume in the annulus isreduced to form a convex surface 412 (shown in phantom) extending fromthe first end 402. It is this convex surface 312 that is observed as thesample within the annulus 402 is set into resonant oscillation as aresult of stimulation of the cartridge 10/200.

While the structures 300 and 400 are shown as circular cylinders, thesample retention structure is not limited in its shape. The sampleretention structure need only have a wall structure, e.g., wall 314 orwall 414 of the respective structures 300 and 300, to contain the samplebut leaving the sample with at least two free surfaces permittingobservable resonant vibration of the sample. For example, the sampleretention structure may be oval, prismatic, conical, or virtually anyother suitable structure and geometric shape that permits excitation ofthe sample to resonant vibration and observation of the motion of thesample in response to the excitation.

FIG. 7 illustrates a cartridge 500, which is similar to cartridge 10 inthat it is configured for a single test although it could be configuredfor multiple tests, such as is the case with cartridge 200. The sampletesting cartridge 500 includes a sample processing portion 512, a sampleretention portion 514 within a card body 516.

The sample-processing portion 512 includes a port 518 through which aliquid sample, such as the previously described sample 100, may beintroduced into the sample-processing portion 512. The port 518 may beself-sealing (as in a septum or other automatic sealing mechanism) suchthat the sample once introduced into the cartridge 500 does not flow,leak, seep, etc. from the cartridge. The port 518 communicates with areservoir 520 into which the sample is initially received. The sampleprocessing portion 512 additionally includes channels, via, wastechambers, passages and similar structures 522; a bellows or pump 524 andvalves 526 to control movement of the sample 100 or a portion thereofthrough the sample processing portion 512 responsive to actuation of thebellows 524 to prepare the sample 100 for testing.

Pneumatic force, which can be applied pressure, drawn vacuum orcombinations thereof, and in a preferred implementation is vacuum, maybe used directly on the sample 100 to move it into the cartridge 500 andto manipulate the various elements of the cartridge 500. In theillustrated implementation, vacuum applied at a central port 519 causesthe sample 100 to load into a staging area 520 and further draws thesample 100 into the bellows 524. The sample 100 is drawn up to thehydrophobic vents 528, allowing careful control of the sample fluidvolumes solely with the card geometry. As with the cartridges 10 or 200,it is unnecessary to monitor loading time or otherwise actively sense ofthe volume of the sample 100, simplifying the structure and operation ofthe cartridge 500. Likewise, the sample is prepared for in a manner asdescribed above and communicated to a sample retention structure 514.

To excite the sample 100 within the cartridge 500, an exciting stimulusmay be applied to the entire cartridge 500. Arrows “A”, “B” or “C”,indicate single or multiple excitation of the cartridge, and hence thesample, rotational excitation of the cartridge or combinations thereof.Excitation of the cartridge 500 results in corresponding excitation ofthe sample 100 to resonance, which in turn is observed in order toderive hemostasis characteristics.

Although certain apparatus constructed in accordance with the teachingsof the invention and methods have been described herein, the scope ofcoverage of this patent is not limited thereto. Generally, apparatus andmethods are provided yielding for the first time continuous, accurateresults starting from liquid blood and following the transition to a gelstate. Observation of the increasing resonant frequency of the sampleprovides a direct measure of the elastic properties of the material andcorresponding hemostasis characteristics.

This patent covers all examples of the teachings of the invention fairlyfalling within the scope of the appended claims either literally orunder the doctrine of equivalents.

We claim:
 1. A method of testing blood hemostasis comprising:introducing a blood sample into a fluid processing structure, the fluidprocessing structure having a portion containing a reagent; processingat least a portion of the blood sample within the fluid processingstructure by contacting the portion with the reagent; communicating theprocessed portion to a sample retention structure portion of the fluidprocessing structure; and testing the processed portion by vibratoryexcitation of the processed portion to obtain data indicative of atleast one hemostasis parameter.
 2. The method of claim 1, whereinreagent is disposed within a passage of the fluid processing structure,and processing comprises passing the portion through the passage.
 3. Themethod of claim 1, wherein one or more reagents is disposed in one ormore fluid communicating portions or passages of the fluid processingstructure, and processing comprises communicating the portion throughthe fluid processing structure.
 4. The method of claim 1, wherein one ormore reagents is disposed in one or more fluid communicating portions orpassages of the fluid processing structure, and processing comprisescontacting the blood sample with a reagent in at least one reagentcontaining portion of the fluid processing structure.
 5. The method ofclaim 1, wherein the fluid processing structure comprises a pump andwherein passing the portion through the passage comprises pumping theportion through the passage.
 6. The method of claim 5, wherein pumpingthe portion through the passage comprises a predetermined flow profile.7. The method of claim 5, wherein the predetermined flow profilecomprises at least one of a step, a ramp and an arc.
 8. The method ofclaim 5, wherein the predetermined flow profile is selected to reduceshearing of the blood sample.
 9. The method of claim 1, whereincommunicating the processed portion to the sample retention apparatuscomprises pumping the sample to the sample retention apparatus.
 10. Themethod of claim 1, the fluid processing structure comprising first andsecond passages, and processing comprises passing a first portion of theblood sample through the first passage and passing a second portion ofthe blood sample through the second passage.
 11. The method of claim 10,comprising communicating the processed first portion to a first sampleretention structure portion and communicating the processed secondportion to a second sample retention structure portion, andsimultaneously testing the processed first portion to obtain a firsthemostasis parameter, and testing the processed second portion to obtaina second hemostasis parameter.
 12. The method of claim 1, whereintesting comprises exciting the processed portion to resonant vibration.13. The method of claim 12, wherein the hemostasis parameter comprisesat least one of: time to initial clot formation, rate of clot formation,maximum clot strength and degree of clot lysis.
 14. The method of claim1, comprising communicating the data via a network.
 15. The method ofclaim 1, comprising graphically depicting the data.
 16. The method ofclaim 1, the fluid processing portion and the sample retention structurebeing portions of a cartridge, and wherein testing the processed portionby vibratory excitation comprises applying an excitation to thecartridge.
 17. The method of claim 1, the fluid processing portion andthe sample retention structure being portions of a cartridge, andwherein testing the processed portion by vibratory excitation comprisesapplying an excitation to a portion of the cartridge containing thesample retention structure.
 18. The method of claim 1, the methodcomprising sizing the sample volume by selectively filling a volumetricportion of the fluid processing portion
 19. The method of claim 1, themethod comprising sizing the sample volume by sizing a geometric volumeof the fluid processing portion.
 20. The method of claim 1, the methodcomprising observing a portion of the processed portion during vibratoryexcitation.